DAB Peroxidase staining (View forum version)



baboon

Posted 19 August 2009 - 11:14 AM

Hi,

I am trying to do IHC using an HRP-labeled secondary antibody. To detect the antibody I am using DAB + Nickel chloride as my substrate.
My question pertains to dissolving DAB substrate. I am trying to dissolve DAB in PBT (0.2% BSA, 0.1% triton X-100 in PBS) but it takes forever to dissolve it. Does anyone know a better way of how to dissolve DAB?

Thx.

pesji

Posted 21 August 2009 - 03:45 AM

Hi,

I am trying to do IHC using an HRP-labeled secondary antibody. To detect the antibody I am using DAB + Nickel chloride as my substrate.
My question pertains to dissolving DAB substrate. I am trying to dissolve DAB in PBT (0.2% BSA, 0.1% triton X-100 in PBS) but it takes forever to dissolve it. Does anyone know a better way of how to dissolve DAB?

Thx.

Check the pH of your pBS it has to be 7.3, I would rather dissolve in pBS only

baboon

Posted 25 August 2009 - 09:46 AM

Thanks,
I shall try dissolving in PBS.....

miraclestrain

Posted 18 October 2009 - 08:13 PM

Hi,

I am trying to do IHC using an HRP-labeled secondary antibody. To detect the antibody I am using DAB + Nickel chloride as my substrate.
My question pertains to dissolving DAB substrate. I am trying to dissolve DAB in PBT (0.2% BSA, 0.1% triton X-100 in PBS) but it takes forever to dissolve it. Does anyone know a better way of how to dissolve DAB?

Thx.



Hi there,

I am using the same stuff, but i am using anti mouse HRP, for the DAB I am following the protocol provided by sigma, which is- Crush DAB- (atleast for 1 hour, to make it as a fine powder), try dissolving it in 3-5 ml warm ethanol, finally disolve it in PBS. I would like to discuss more about this, i am from Melbourne and you are?

MS